长链非编码 RNA DANCR 通过调节 FOXO1 表达抑制成骨细胞分化从而参与全髋关节置换术后溶骨。

LncRNA DANCR involved osteolysis after total hip arthroplasty by regulating FOXO1 expression to inhibit osteoblast differentiation.

机构信息

Department of Articular Orthopaedics, Changzhou First People's Hospital, The Third Affiliated Hospital of Soochow University, No.185 Juqian Rd, Changzhou, Jiangsu, 213003, China.

出版信息

J Biomed Sci. 2018 Jan 16;25(1):4. doi: 10.1186/s12929-018-0406-8.

BACKGROUND

Aseptic loosening of artificial hip joint is a major complication affecting the long-term use of the artificial hip joint, and is the main cause of joint replacement failure. However, the mechanism of aseptic loosening of THR has not yet cleared. The aim of this study was to investigate the underlying mechanism of DANCR in osteoblast differentiation (OD).

METHODS

We detected the expressions of DANCR and FOXO1 in clinical samples and mesenchymal stem cells (MSCs) by qRT-PCR and western blotting. The effects of polymethylmethacrylate (PMMA) on OD of MSCs were examined by alkaline phosphatase (ALP) activity and Alizarin Red S (ARS) staining. The expressions of OD markers were measured by qRT-PCR and western blotting. The mechanism of DANCR in OD was detected by RNA pull-down, RNA immunoprecipitation (RIP) assay and ubiquitination assays.

RESULTS

Compared with the surrounding normal tissues, DANCR expression was up-regulated and FOXO1 expression was down-regulated in periprosthetic tissues. PMMA suppressed ALP activity, increased DANCR expression, and decreased the expressions of FOXO1, Runx2, Osterix (Ostx) and osteocalcin (OCN). ARS staining showed that PMMA inhibited the OD of MSCs. Knockdown of DANCR attenuated the inhibitory effect of PMMA on OD. Knockdown of FOXO1 could reverse the effect of si-DANC. RNA pull-down and RIP assay implicated that DANCR bound to FOXO1. Ubiquitination assay indicated that si-DANCR could repress Skp2-mediated ubiquitination of FOXO1.

CONCLUSION

LncRNA DANCR could inhibit OD by regulating FOXO1 expression.

背景

人工髋关节无菌性松动是影响人工髋关节长期使用的主要并发症，也是关节置换失败的主要原因。然而，THR 无菌性松动的机制尚未明确。本研究旨在探讨 DANCR 在成骨细胞分化（OD）中的潜在作用机制。

方法

我们通过 qRT-PCR 和 Western blot 检测了临床样本和间充质干细胞（MSCs）中 DANCR 和 FOXO1 的表达。通过碱性磷酸酶（ALP）活性和茜素红 S（ARS）染色检测聚甲基丙烯酸甲酯（PMMA）对 MSCs OD 的影响。通过 qRT-PCR 和 Western blot 检测 OD 标志物的表达。通过 RNA 下拉、RNA 免疫沉淀（RIP）实验和泛素化实验检测 DANCR 在 OD 中的作用机制。

结果

与周围正常组织相比，假体周围组织中 DANCR 的表达上调，FOXO1 的表达下调。PMMA 抑制 ALP 活性，增加 DANCR 的表达，降低 FOXO1、Runx2、Osterix（Ostx）和骨钙素（OCN）的表达。ARS 染色表明 PMMA 抑制 MSCs 的 OD。DANCR 敲低减弱了 PMMA 对 OD 的抑制作用。si-DANC 的敲低可以逆转 si-DANC 的作用。RNA 下拉和 RIP 实验表明 DANCR 与 FOXO1 结合。泛素化实验表明 si-DANCR 可以抑制 Skp2 介导的 FOXO1 泛素化。